Adenovirus-mediated gene therapy of osteoblasts in vitro and in vivo

Modulation of biological pathways governing osteogenesis may accelerate oss eous regeneration and reduce the incidence of complications associated with fracture healing. Transforming growth factor beta 1 (TGF-beta 1) is a pote nt growth factor implicated in the regulation of osteogenesis and fracture repair. The use of recombinant proteins, however, has significant disadvant ages and has limited the clinical utility of these molecules. Targeted gene therapy using adenovirus vectors is a technique that may circumvent diffic ulties associated with growth factor delivery. In this study, we investigat e the efficacy of replication-deficient adenoviruses containing the human T GF-beta 1 and the bacterial lacZ genes in transfecting osteoblasts in vitro and osseous tissues in vivo. We demonstrate that adenovirus-mediated gene therapy efficiently transfects osteoblasts in vitro with the TGF-beta 1 vir us causing a marked up-regulation in TGF-beta 1 mRNA expression even 7 days after transfection. Increased TGF-beta 1 mRNA expression was efficiently t ranslated into protein production and resulted in approximately a 46-fold i ncrease in TGF-beta 1 synthesis as compared with control cells (vehicle- or B-galactosidase-transfected). Moreover. virally produced TGF-beta 1 was fu nctionally active and regulated the expression of collagen I alpha I (5-fol d increase) and the vascular endothelial growth factor (2.5-fold increase). Using an adenovirus vector encoding the Escherichia coli LacZ gene, we dem onstrated that adenovirus-mediated gene transfer efficiently transfects ost eoblasts and osteocytes in vivo and that transfection can be performed by a simple percutaneous injection. Finally, we show that delivery of the hTGF- beta 1 gene to osseous tissues in vivo results in significant changes in th e epiphyseal plate primarily as a result of increased thickness of the prov isional calcification zone.